In patients with diabetes, obesity, or cardiovascular disease, the circulating proinflammatory mediators are often elevated. We made the novel observation that intestinal mucosal mast cells (MMC) are activated by fat absorption. In our well established conscious lymph fistula rats, we showed that fat absorption activates the intestinal mucosal mast cells (MMC) and they degranulate as evidenced by the marked increase (~ 20 fold) of the MMC marker protease II in intestinal lymph relative to fasting lymph (pre-ingestion of fat). This observation is very exciting from the standpoint of gut physiology; it may have profound clinical implications. We consume fat a few times a day and so the MMC are activated frequently during the day. With the activation and degranulation of MMC, the gut may be an important contributor to the circulating proinflammatory mediators. We hypothesized that the absorption of lipids and the formation and secretion of chylomicrons (CMs) activate the MMCs. This action is lipid specific and is not shared by proteins or carbohydrates. We further hypothesize that acute and chronic consumption of fat, the type of fat, influence its ability to activate the MMCs and that the order of potency is omega 6 (linoleate & arachidonate) > omega 9 (oleate) > omega 3 fatty acids (linolenate). To test these hypotheses, we proposed the following studies: Specific Aim 1. We will determine the relationship between intestinal fat absorption and MMC activation. Subaims: 1) to further characterize the activation of MMC by measuring the release of mast cell mediators in lymph and tissue during fat absorption, we will determine if the release of lipid mediators, cytokines, chemokines is mostly from MMC by using the mast cell deficient Ws/Ws rats; 2) to determine if the presence or absence of MMC stabilizers affects intestinal uptake and lymphatic transport of dietary lipids. We will also study fat absorption in Ws/Ws rats. Specific Aim 2. We hypothesize that the production of CMs, resulting from dietary polyunsaturated long-chain fatty acids (FA) linoleic acid (18:2, n-6) is a potent determinant of the mast cell activation and this action is differently affected by the type of fatty acids. We will determine the acute and chronic exposure to fatty acids with different degree of unsaturation and the type (n-3, n-6, and n-9) fatty acids on MMC activation by fat absorption. Specific Aim 3. We will test our hypothesis that fatty acid or the processing of CMs by the enterocytes results in the release of factor/factors that cause the activation of the MMCs (e.g. NF-kB response). To test our hypotheses, we have proposed the following 2 subaims. 1) We will study the in vitro effects of different types of FAs on rat basophil leukemia cells (RBL-2H3 cells, a surrogate of the rat MMC) activation through NF-kB signaling pathway; 2) We will study the interaction of chylous lymph and its components e.g. chylomicrons (CMs) or the media containing CMs by the intestinal epithelial cell line (Caco-2 cells) on RBC-2H3 cell NF-kB response. If the outcome is positive, we will isolate the factor/factors involved. PUBLIC HEALTH RELEVANCE: By focusing on the relationship between fat absorption and mucosal mast cell degranulation, our proposed studies will identify the role of gut-derived inflammatory factors in the normal process of dietary fat absorption and transport, identify which inflammatory agents are generated by normal fat absorption and may enter the general circulation, and contribute to the etiology of various diseases such as diabetes, metabolic syndrome, and atherosclerosis and coronary heart disease. Our results will also greatly enhance our understanding of function of gastrointestinal lymphatics in health and disease by clarifying the changes which occur in GI lymph parameters during normal physiological states such as fasting and lipid absorption, laying the ground work for future studies comparing the differences in lymph between diseased states and normal physiological states.